chip grade antibodies anti h3k4me3  (Cell Signaling Technology Inc)

Journal: Life Science Alliance

Article Title: Dectin-1 epigenetic reprogramming rescues senescent-like Treg function in allergic asthma

doi: 10.26508/lsa.202503552

Figure Lengend Snippet: (A) Binding of KQS-1 to purified human Tregs. Cells were incubated with KQS-1 and analyzed by flow cytometry to determine the proportion of KQS-1–positive cells. Representative gating strategies are shown in . (B, C, D) Raf-1 and ROS dependence of KQS-1–primed Treg responses. Tregs were treated with KQS-1 alone or in combination with the Raf-1 inhibitor GW5074 (10 μM) or the ROS scavenger N-acetylcysteine (NAC, 5 mM). (B) H3K4me3 enrichment at the FOXP3 promoter assessed by ChIP–qPCR and expressed in arbitrary units (a.u.). (C) In vitro suppressive capacity of treated Tregs. The dashed line indicates the mean suppression achieved by KQS-1–treated cells. Representative flow cytometry plots are provided in . (D) TGF-β protein levels in culture supernatants under the indicated conditions. (C) Data are presented as the mean ± SD, with each dot representing an individual donor (n = 10 biologically independent replicates for panel (C)). Statistical significance was determined by one-way ANOVA followed by Tukey’s multiple comparisons test. **** P < 0.0001.

Article Snippet: ChIP-grade antibodies anti-H3K4me3 (clone: C42D8) and anti-H3K27ac (clone: D8L3M) (Cell Signaling Technology); ChIP assay kit (Millipore); and Illumina Infinium HumanMethylation450K BeadChip (Illumina) were used to perform ChIP–qPCR to analyze epigenetic changes at specific genes.

Techniques: Binding Assay, Purification, Incubation, Flow Cytometry, ChIP-qPCR, In Vitro

Journal: Life Science Alliance

Article Title: Dectin-1 epigenetic reprogramming rescues senescent-like Treg function in allergic asthma

doi: 10.26508/lsa.202503552

Figure Lengend Snippet: (A) ChIP–qPCR analysis of active histone marks at key Treg signature loci. Enrichment of H3K4me3 at the FOXP3 promoter and IL-10 locus, as well as global H3K27ac levels, was assessed in KQS-1–treated and control cells. (B) ATAC-seq volcano plot comparing chromatin accessibility in KQS-1–treated versus untreated Tregs, highlighting newly accessible genomic regions. (C) Genomic annotation of KQS-1–induced accessible regions, categorized as promoters (±3 kb of the transcription start site), enhancers (distal regulatory elements), or intergenic regions. Distribution of accessibility changes across genomic features is shown. (D) Chromatin accessibility at IL-10 and FOXP3 promoter regions. Upper panels show representative ATAC-seq signal tracks; lower panels present quantification of accessibility changes after KQS-1 treatment. (E) Gene ontology (GO) enrichment analysis of genes proximal to KQS-1–induced accessible regions, illustrating enrichment of immune regulatory and T cell–associated pathways. (F) DNA methylation profiling of regulatory regions using the Illumina 450K array. Lollipop plots depict changes in methylation (Δβ) between KQS-1–treated and untreated Tregs, highlighting focal hypomethylation at FOXP3 and IL-10 regulatory CpG sites. ChIP–qPCR data are presented as the mean ± SD (n = 8 biological replicates), with individual points representing replicates. ATAC-seq analysis identified 1,306 newly accessible regions (FDR < 0.05). For genomic distribution analysis, box plots show median and interquartile range (IQR), with individual dots representing peaks. DNA methylation data are shown as mean Δβ with 95% confidence intervals. Statistical significance was determined using paired or unpaired t tests, one-way ANOVA with Tukey’s post hoc test, and the Bonferroni correction for multiple comparisons, as appropriate. ** P < 0.01, *** P < 0.001, **** P < 0.0001; ns, not significant.

Article Snippet: ChIP-grade antibodies anti-H3K4me3 (clone: C42D8) and anti-H3K27ac (clone: D8L3M) (Cell Signaling Technology); ChIP assay kit (Millipore); and Illumina Infinium HumanMethylation450K BeadChip (Illumina) were used to perform ChIP–qPCR to analyze epigenetic changes at specific genes.

Techniques: ChIP-qPCR, Control, DNA Methylation Assay, Methylation

Journal: Life Science Alliance

Article Title: Dectin-1 epigenetic reprogramming rescues senescent-like Treg function in allergic asthma

doi: 10.26508/lsa.202503552

Figure Lengend Snippet: Gain-of-function analysis in Dectin-1-low Tregs after lentiviral transduction with Dectin-1 (Dectin-1 OE) or empty vector control. (A, B) Flow cytometry analysis showing a significant increase in KQS-1 binding (MFI) in Dectin-1 OE cells compared with controls (A), accompanied by restored H3K4me3 epigenetic marks at the FOXP3 promoter region as measured by ChIP–qPCR (B). (C) Suppressive efficiency of Tregs against responder T-cell proliferation, demonstrating enhanced inhibitory capacity upon Dectin-1 restoration. Data are presented as box-and-whisker plots with individual samples shown as dots (n = 5 per group). Statistical significance was determined by an unpaired t test; * P < 0.05, ** P < 0.01.

Article Snippet: ChIP-grade antibodies anti-H3K4me3 (clone: C42D8) and anti-H3K27ac (clone: D8L3M) (Cell Signaling Technology); ChIP assay kit (Millipore); and Illumina Infinium HumanMethylation450K BeadChip (Illumina) were used to perform ChIP–qPCR to analyze epigenetic changes at specific genes.

Techniques: Transduction, Plasmid Preparation, Control, Flow Cytometry, Binding Assay, ChIP-qPCR, Whisker Assay

Journal: Sports Medicine and Health Science

Article Title: Exercise preconditioning prevents immobilization-induced skeletal muscle atrophy by activating Prmt1-p38/ATF2-Sesn1 signaling axis in C57BL/6J mice

doi: 10.1016/j.smhs.2025.04.001

Figure Lengend Snippet: 10-week combined exercise preconditioning prevented 1-week hindlimb immobilization-induced muscle atrophy in mice. (A–C) Body weight, gross weight gain and food intake data during 10-week exercise preconditioning. ∗ p ​< ​0.05 vs. C; # p ​< ​0.05 vs. E. Two-way ANOVA was used and data are shown as means ​± ​standard error of the mean ( SEM ) (C, n ​= ​16, E, n ​= ​8, E+5003, n ​= ​8). (D) Food intake data during 1-week immobilization. ∗ p ​< ​0.05 vs. C. Two-way ANOVA was used and data are shown as means ​± ​ SEM . (E–F) aDMA level indicated for Prmt1 enzymatic activity after 10-week TC-E−5003 administration via subcutaneous injection at a dose of 2 ​mg/kg body weight, once daily, 5 days a week. (G–J) Skeletal muscle functions tests, G, grip strength, H, suspension time of hang test, I, time of the latency to fall in rotarod test, J, maximum voluntary climbing capacity (MVCC) test. (K–P) Total hindlimb mass and specific muscle mass of various parts of hindlimb, GAS, gastrocnemius, SOL, soleus, QUA, quadriceps femoris, lateralis, TA, tibialis anterior, EDL, extensor digitorum longus. H-S, ∗ p ​< ​0.05 vs. C; ∗∗ p ​< ​0.01 vs. C; # p ​< ​0.05 vs. Im; ## p ​< ​0.01 vs. Im; $ p ​< ​0.05 vs. E ​+ ​Im; $$ p ​< ​0.01 vs. E ​+ ​Im. Two-way ANOVA was used and data are shown as means ​± ​ SEM (C, n ​= ​8, Im, n ​= ​8, E ​+ ​Im, n ​= ​8, E+5003+Im, n ​= ​8).

Article Snippet: ChIP grade antibodies to Prmt1 (1:100, Proteintech, USA) were used to perform immunoprecipitation.

Techniques: Activity Assay, Injection, Suspension

Journal: Sports Medicine and Health Science

Article Title: Exercise preconditioning prevents immobilization-induced skeletal muscle atrophy by activating Prmt1-p38/ATF2-Sesn1 signaling axis in C57BL/6J mice

doi: 10.1016/j.smhs.2025.04.001

Figure Lengend Snippet: 10-week combined exercise preconditioning ameliorated 1-week hindlimb immobilization-induced imbalance between synthesis and degradation of protein in skeletal muscle. (A) Representative images of hematoxylin and eosin staining (H&E) of GAS muscle cross-sections. Scale bar ​= ​50 ​μm. (B) The average CSA of GAS muscle was quantified. (C–D) Real-time PCR results of TIRM63 (MuRF1), FBXO32 (Atrogin-1), Prmt1, and Sesn1 in GAS muscle. (E–N) Western blot results of Prmt1, Sesn1, p-FoxO3a, FoxO3a, Atrogin-1, MuRF1, IGF-1, pAkt-S473, Akt, mTOR, Raptor in GAS muscle. B-N, ∗ p ​< ​0.05 vs. C; ∗∗ p ​< ​0.01 vs. C; # p ​< ​0.05 vs. Im; ## p ​< ​0.01 vs. Im; $ p ​< ​0.05 vs. E ​+ ​Im; $$ p ​< ​0.01 vs. E ​+ ​Im. Two-way ANOVA was used and data are shown as means ​± ​standard error of the mean (C, n ​= ​8, Im, n ​= ​8, E ​+ ​Im, n ​= ​8, E+5003+Im, n ​= ​8).

Article Snippet: ChIP grade antibodies to Prmt1 (1:100, Proteintech, USA) were used to perform immunoprecipitation.

Techniques: Staining, Real-time Polymerase Chain Reaction, Western Blot

Journal: Sports Medicine and Health Science

Article Title: Exercise preconditioning prevents immobilization-induced skeletal muscle atrophy by activating Prmt1-p38/ATF2-Sesn1 signaling axis in C57BL/6J mice

doi: 10.1016/j.smhs.2025.04.001

Figure Lengend Snippet: Exercise preconditioning increased Sestrin1 (Sesn1) expression by activating protein arginine methyltransferase 1 (Prmt1)-p38/activating transcription factor 2 (ATF2) signaling in mice. (A) ChIP analysis of Prmt1 binding to the ATF2 sequence of mouse Sesn1 promoter in GAS muscle. (B–D) Western blot results of p-p38-T180/182, p38, and ATF2 in GAS muscle. (E–F) Co-IP results, IP: Prmt1, GAS muscle was used. (G–H) Co-IP results, IP: ATF2, GAS muscle was used. (I–J) Co-IP results, IP: p38, GAS muscle was used. A-J, ∗ p ​< ​0.05 vs. C; ∗∗ p ​< ​0.01 vs. C; # p ​< ​0.05 vs. Im; ## p ​< ​0.01 vs. Im; $ p ​< ​0.05 vs. E ​+ ​Im; $$ p ​< ​0.01 vs. E ​+ ​Im. Two-way ANOVA was used and data are shown as means ​± ​standard error of the mean (C, n ​= ​8, Im, n ​= ​8, E ​+ ​Im, n ​= ​8, E+5003+Im, n ​= ​8).

Article Snippet: ChIP grade antibodies to Prmt1 (1:100, Proteintech, USA) were used to perform immunoprecipitation.

Techniques: Expressing, Binding Assay, Sequencing, Western Blot, Co-Immunoprecipitation Assay

Journal: Sports Medicine and Health Science

Article Title: Exercise preconditioning prevents immobilization-induced skeletal muscle atrophy by activating Prmt1-p38/ATF2-Sesn1 signaling axis in C57BL/6J mice

doi: 10.1016/j.smhs.2025.04.001

Figure Lengend Snippet: Protein arginine methyltransferase 1 (Prmt1)-Sestrin1 (Sesn1) coordinated the differentiation of C2C12 myoblasts into mature myotubes. (A–B) H&E staining of C2C12 myotubes at the 7 th day of differentiation with si-Sesn, and Ad-Sesn1 treatment. Scale bar ​= ​100 ​μm. (C–D) Western blot results of Sesn1, FoxO3a, Atrogin-1 and MuRF1 in C2C12. A-D, C, control, si-Sesn1, si-RNA of Sesn1, Ad-Sesn1, Adenoviruses overexpressing Sesn1, ∗ p ​< ​0.05 vs. C; ∗∗ p ​< ​0.01 vs. C; # p ​< ​0.05 vs. si-Sesn1; ## p ​< ​0.01 vs. Si-Sesn1. Two-way ANOVA was used and data are shown as means ​± ​standard error of the mean ( SEM ) ( n ​= ​6 in each group). (E–F) H&E staining of C2C12 myotubes at the 7 th day of differentiation with si-Prmt1, and Ad-Prmt1. Scale bar ​= ​100 ​μm. (G–N) Western blot results of Prmt1, p-p38-T180/182, p38, ATF2, Sesn1, FoxO3a, Atrogin-1 and MuRF1 in C2C12. E-N, C, control, si-Prmt1, si-RNA of Prmt1, Ad-Prmt1, Adenoviruses overexpressing Prmt1, ∗ p ​< ​0.05 vs. C; ∗∗ p ​< ​0.01 vs. C; # p ​< ​0.05 vs. si-Prmt1; ## p ​< ​0.01 vs. si-Prmt1. Two-way ANOVA was used and data are shown as means ​± ​ SEM ( n ​= ​3 in each group). (O–R) Western blot results of Prmt1, ATF2 and Sesn1 in C2C12 myoblasts, ∗ p ​< ​0.05; ∗∗ p ​< ​0.01, unpaired Student's t -test was used and data are shown as means ​± ​ SEM ( n ​= ​6 in each group). (S–W) Western blot results of Prmt1, FoxO3a, Atrogin-1 and MuRF1 in C2C12 myoblasts, ∗ p ​< ​0.05; ∗∗ p ​< ​0.01, unpaired Student's t -test was used and data are shown as means ​± ​ SEM ( n ​= ​6 in each group).

Article Snippet: ChIP grade antibodies to Prmt1 (1:100, Proteintech, USA) were used to perform immunoprecipitation.

Techniques: Staining, Western Blot, Control

Journal: Sports Medicine and Health Science

Article Title: Exercise preconditioning prevents immobilization-induced skeletal muscle atrophy by activating Prmt1-p38/ATF2-Sesn1 signaling axis in C57BL/6J mice

doi: 10.1016/j.smhs.2025.04.001

Figure Lengend Snippet: Exercise preconditioning activated AMP-Activated protein kinase α2 (AMPKα2)-transcriptional co-activator PPAR-γ co-activator-1 α (PGC-1α) through protein arginine methyltransferase 1 (Prmt1)-Sestrin1 (Sesn1) in immobilized mice. (A–C) Western blot results of p-AMPKα2, AMPKα2, and PGC-1α in GAS muscle. (D–E) Co-IP results, IP: Prmt1, GAS muscle was used. (F–G) Co-IP results, IP: PGC-1α, GAS muscle was used. (H–I) Co-IP results, IP: Sesn1, GAS muscle was used. (J–K) Co-IP results, IP: PGC-1α, GAS muscle was used. (L–M) Co-IP results, IP: AMPKα2, GAS muscle was used. A-M, ∗ p ​< ​0.05 vs. C; ∗∗ p ​< ​0.01 vs. C; # p ​< ​0.05 vs. Im; ## p ​< ​0.01 vs. Im; $ p ​< ​0.05 vs. E ​+ ​Im; $$ p ​< ​0.01 vs. E ​+ ​Im. Two-way ANOVA was used and data are shown as means ​± ​standard error of the mean (C, n ​= ​8, Im, n ​= ​8, E ​+ ​Im, n ​= ​8, E+5003+Im, n ​= ​8).

Article Snippet: ChIP grade antibodies to Prmt1 (1:100, Proteintech, USA) were used to perform immunoprecipitation.

Techniques: Western Blot, Co-Immunoprecipitation Assay

Journal: Sports Medicine and Health Science

Article Title: Exercise preconditioning prevents immobilization-induced skeletal muscle atrophy by activating Prmt1-p38/ATF2-Sesn1 signaling axis in C57BL/6J mice

doi: 10.1016/j.smhs.2025.04.001

Figure Lengend Snippet: Exercise preconditioning prevented muscle atrophy through protein arginine methyltransferase 1 (Prmt1)-Sestrin1 (Sesn1)-transcriptional co-activator PPAR-γ co-activator-1 α (PGC-1α)-mediated skeletal muscle regeneration. (A–B) Western blot results of MyoD, Myf5, MEF2 and MyoG in GAS muscle. (C–D) Co-IP results, IP: MyoD, GAS muscle was used. A-B, ∗ p ​< ​0.05 vs. C; ∗∗ p ​< ​0.01 vs. C; ## p ​< ​0.01 vs. Im; $ p ​< ​0.05 vs. E ​+ ​Im; $$ p ​< ​0.01 vs. E ​+ ​Im. Two-way ANOVA was used, and data are shown as means ​± ​standard error of the mean ( SEM ) (C, n ​= ​8, Im, n ​= ​8, E ​+ ​Im, n ​= ​8, E+5003+Im, n ​= ​8). (E–F) H&E staining of myotubes at each differentiation time points of C2C12 myoblasts overexpressing Sesn1. Scale bar ​= ​100 ​μm ∗ p ​< ​0.05 vs. Ad-Sesn1-, unpaired Student's t-test was used and data are shown as means ​± ​ SEM ( n ​= ​3 in each group). (G–H) Western blot results of Sesn1, Prmt1, PGC-1α, MyoD, MEF2, Myf5, and MyoG in C2C12 myoblasts at each time points of differentiation. ∗ p ​< ​0.05 vs. D0, unpaired Student's t -test was used and data are shown as means ​± ​ SEM ( n ​= ​6 in each group). (I–J) Western blot results of PGC-1α in nucleus of C2C12 myoblasts overexpressing Sesn1 at each time points of differentiation. ∗∗ p ​< ​0.01 vs. Ad-Sesn1, unpaired Student's t -test was used and data are shown as means ​± ​ SEM ( n ​= ​6 in each group). (K–L) Western blot results of PGC-1α in cytoplasm of C2C12 myoblasts overexpressing Sesn1 at each time points of differentiation. ∗∗ p ​< ​0.01 vs. Ad-Sesn1-, unpaired Student's t -test was used and data are shown as means ​± ​ SEM ( n ​= ​6 in each group).

Article Snippet: ChIP grade antibodies to Prmt1 (1:100, Proteintech, USA) were used to perform immunoprecipitation.

Techniques: Western Blot, Co-Immunoprecipitation Assay, Staining